Protein interactions that regulate cell polarity

描述(申请人提供)：这项工作的长期目标是了解细胞极化的分子基础。极性是细胞的基本属性，对正常发育和成人生理都是必需的。例如，在发育过程中，细胞命运决定因素在分裂细胞时两极分化，作为产生细胞类型多样性的机制，细胞极性的丧失是包括癌症在内的许多疾病状态的标志。为了在空间和时间上精确地确定细胞的极性，必须解释细胞信号，并最终将其与相关细胞成分的分离相联系。在不同类型的细胞中，极性由进化上保守的PAR复合体控制，该复合体由Bazooka(BAZ；又名PAR-3)、PAR-6和非典型蛋白激酶C(APKC)组成。Rho GTPase CDc42与PAR-6相互作用，是PAR复合体靶向和激活的主要决定因素。该提案的第一个具体目标是确定当cdc42与PAR-6结合时，如何通过激活aPKC激酶来改变PAR复合体的活性。我们正在检验一种假设，即aPKC调控源于存在于PAR-6中的PDZ蛋白相互作用结构域的激活。为了实现这一目标，将采用生化、细胞生物学和结构学相结合的方法。第二个目标是确定在特定细胞位置引起激活的CDC42的分子途径。我们正在测试一种假设，即核苷酸交换因子激活细胞皮质特定区域的CDC42，而其他因素抑制其他区域的激活。为此，我们将利用神经母细胞作为模型系统来揭示调节CDC42空间和时间激活的分子通路，这最终是PAR复合体靶向的责任。为实现这一目标，将采用遗传和生化相结合的方法。最后，我们正在研究aPKC激酶活性如何与神经母细胞中细胞命运决定因素的不对称分离相耦合，并假设这一过程需要aPKC直接磷酸化碱基分离蛋白Miranda。了解导致PAR复合体耦合招募和激活的分子事件将对细胞极性产生新的见解。公共卫生关注性我们身体中的许多细胞，如为环境提供物理屏障的皮肤细胞，都是极化的，极性丧失是包括癌症在内的许多疾病的标志。在这项工作中，我们正在研究一组被称为PAR复合体的三种蛋白质，它们调节正常发育和成人生理所需的细胞极性。由于极性的丧失与人类疾病有关，提高我们对控制这一过程的分子的理解将有助于我们对疾病状态机制的了解。

DESCRIPTION (provided by applicant): The long-term objective of this work is to understand the molecular basis by which cells are polarized. Polarity is a fundamental property of cells that is required for proper development as well as adult physiology. For example, during development cell fate determinants are polarized in dividing cells as a mechanism for generating cell type diversity and the loss of cell polarity is a hallmark of many disease states, including cancer. For spatially and temporally precise establishment of cell polarity to occur, cellular signals must be interpreted and ultimately coupled to the segregation of the relevant cellular components. In diverse cell types, polarity is controlled by the evolutionarily conserved Par complex consisting of Bazooka (Baz; aka Par-3), Par-6, and atypical Protein Kinase C (aPKC). The Rho GTPase Cdc42, which interacts with Par-6, is a primary determinant of Par complex targeting and activation. The first specific aim of the proposal is to determine how Cdc42 alters Par complex activity, by activating the aPKC kinase when it binds to Par-6. We are testing the hypothesis that aPKC regulation arises from activation of a PDZ protein interaction domain present in Par-6. A combined biochemical, cell biological, and structural approach will be utilized for this aim. The second aim is to identify the molecular pathways that give rise to activated Cdc42 at specific cellular sites. We are testing the hypothesis that nucleotide exchange factors activate Cdc42 a certain areas of the cell cortex, and other factors inhibit activation elsewhere. For this aim we will utilize the neuroblast as a model system to uncover the molecular pathways that regulate Cdc42 spatial and temporal activation, which is ultimately responsible for Par complex targeting. A combined genetic and biochemical approach will be utilized for this aim. Finally, we are examining how aPKC kinase activity is coupled to the asymmetric segregation of cell fate determinants in neuroblasts with the hypothesis that direct phosphorylation of the basally segregated protein Miranda by aPKC is required for this process. Understanding the molecular events that lead to the coupled recruitment and activation of the Par complex will yield new insight into cell polarity. PUBLIC HEALTH RELEVANCE Many cells in our body, such as skin cells that provide a physical barrier to the environment, are polarized and loss of polarity is a hallmark of many diseases, including cancer. In this work, we are investigating a set of three proteins, known as the Par complex, that regulate cellular polarities required for proper development and adult physiology. As the loss of polarity is associated with human disease, improving our understanding of the molecules that control this process will contribute to our knowledge of the mechanisms of disease states.